Our long-term objective is to identify gene mutations that cause obesity in humans and to understand their pathogenic effects. This proposal focuses on the G-protein coupled Melanocortin-4 Receptor (MC4R), a gene expressed in the central nervous system and implicated in the regulation of food intake. We had initially reported the first case of human obesity linked to a mutation in the gene encoding the MC4R. In the previous funding period we have extensively studied the prevalence of MC4R mutations in both severely obese children and adult populations; we have comprehensively studied the functional defects of over 50 different obesity causing MC4R mutations; we have further characterized the phenotype of MC4R mutation carriers and have started to study the genotype-phenotype relationship within this disease. Our new specific aims are: 1) To extend the study of the role of the MC4R locus in energy homeostasis in humans. We will extend our genetic studies to the entire MC4R locus by testing for an association between genetic variations outside of the transcriptional unit and minimal promoter of MC4R with obesity related phenotypes in both a large case- control study and a well phenotyped multi-ethinic cohort of 3,075 individuals. We will especially concentrate our efforts on evolutionary conserved regions at the MC4R locus. 2) To extend the study of the genotype-phenotype relationship in obese adult MC4R mutation carriers to the outcome after bariatric surgery. A major goal of identifying genes in which mutations are responsible for common human obesity is to provide more rational approaches to therapy, eventually by stratifying patients into groups in which the effectiveness of different treatments can be determined empirically. Bariatric surgery is currently the most effective therapy for morbid obesity. We will use a large prospectively collected cohort of 2400 patients undergoing bariatric surgery to determine if MC4R mutation carriers have a different outcome than non-carriers. We will also systematically study the functional defects caused by novel mutations detected in this cohort and test if the outcome after bariatric surgery can be correlated to the functional characteristics of the mutations within the group of MC4R mutation carriers. 3) To examine the molecular mechanisms involved in the constitutive activity of the melanocortin-4 receptor by its N-terminal Domain. Through the systematic study of obesity-associated mutations in the N-terminal domain of MC4R we have recently demonstrated that this domain maintains the constitutive activity of MC4R and that this constitutive activity is physiologically relevant. We will further identify the structural features contributing to the constitutive activation the receptor by its N-terminal domain. These experiments could lead to new strategies to pharmacologically target MC4R for the treatment of obesity.